The DNA of eukaryotic organisms is packaged with core histone octamers, linker histones, and other chromosomal proteins to form chromatin fibers. Higher order folding and compaction of chromatin fibers into interphase chromosomes involves both intrinsic and protein-mediated processes. Intrinsic condensation is mediated by the core histone N-terminal domains and the linker histone C-terminal domain. Protein mediated fiber condensation occurs when certain nucleosome binding proteins interact with chromatin fibers and mediate assembly of higher order chromatin suprastructures. In addition to compacting chromosomal DNA, chromatin condensation has been linked to regulation of genomic functions; e.g., transcription. In the present proposal, I plan to use quantitative biochemical and biophysical approaches to characterize the mechanisms of both intrinsic and protein-mediated chromatin fiber condensation, and to determine the structural features of functional genomic elements assembled into chromatin in vivo. Specifically, I propose to: (1) characterize the effects of specific recombinant core and linker histone mutants on intrinsic chromatin fiber condensation, (2) characterize the human transcriptional repressor, MeCP2, as a prototype chromatin architectural protein, and (3) use novel electrophoretic techniques to probe the higher order chromatin structure of the host cell-integrated HTLV-1 viral genome. The proposed studies will be the first to characterize the mechanisms of intrinsic and protein-mediated condensation of biochemically defined model chromatin fibers assembled entirely from recombinant histone and non-histone components. Collectively, the information obtained from these studies will provide a framework for understanding how chromatin fiber condensation is linked to regulation of genomic function. Together, the proposed experiments represent an innovative approach to understanding the higher order structure and dynamics of the eukaryotic genome, and its relation to nuclear function. [unreadable] [unreadable]